ABSTRACT Pancreaticcancerisoneofthemostlethalcancersamongallsolidtumors,withanextremelypoorprognosis, andadismalfive-yearsurvivalrateof7%.Newinsightsintotheregulationofimmuneresponsesandthe malignantprocesshaveledtotheemergenceofnewimmunotherapeuticstrategiestotreatpancreaticcancer. Thechallengesinthedevelopmentoftumorvaccineslieintheidentificationoftumor-associatedantigens, inductionofantigen-specificcellmediatedimmuneresponses,overcomingimmunetolerance,andthe immunosuppressivetumorenvironment.InthisU01application,weproposetodevelopnovelmucin-containing nanovaccineplatformsandcombinethemwithcheckpointblockadeagentsthatcansimultaneouslyinduce long-livedcytotoxicTlymphocyteresponsesandactivateinnateimmunityandmaintaintheTcellresponse, thusovercomingtheimmunosuppresiveenvironment.Amongthemanystructuralandfunctional transformationsthatoccurduringoncogenesis,alteredexpressionofcellsurfaceglycoproteins,suchas mucins,presentsanopportunityforthedevelopmentofvaccinestrategies.WeproposetoutilizeMUC4mucin forimmunotherapeuticnano-formulationsbecausethisproteinisoverexpressedin>90%ofpancreatictumors andundetectableinnormalpancreas(unlikethemostexploredmucinvaccinecandidate,MUC1).Our proposednanoadjuvantplatformconsistsofamphiphilicpolyanhydrideand/orpolyesternanoparticles, containingCpG,andisideallysuitedforprotein-basedsubunitvaccines.Ourcentralhypothesis,basedon significantpeer-reviewedpreliminarydata,isthataleadnanovaccinethatinducesMUC4-specific,cytotoxic CD8+Tcellswilltherapeutically(i.e.,inthepresenceoftumor)provideanti-tumorbenefitsincombinationwith checkpointinhibitors.Wewillpositionthisnanovaccineforpreclinicalstudiesthatwilladvancethe developmentofpancreaticcancervaccinetechnologiesbyaccomplishingthefollowingSpecificAims,eachof whichisboundedbymilestonesandfallbackpositions:Aim1:Formulation,optimization,andimmunological characterizationofMUC4-specificimmuneresponseswithnano-adjuvants.Aim2:Evaluationoflead nanovaccine(s)andcheckpointblockadeagentsinasyngeneicmurinemodelofpancreaticcancer.Aim3: Evaluationofleadnanovaccineandcheckpointblockadeagentsintransgenicmousemodels.Attheendofthe projectperiod,wewilldeliveranovelnanovaccineasaneffectivetherapyforpancreaticcancerpatients. OveralltheproposedstudieswillestablishtheutilityofMUC4,whichisthemostdifferentiallyoverexpressed mucinasatargetforpancreaticcancerimmunotherapy,andthenanotechnologyandcellandanimalmodels generatedinthisprojectwillhavebroaderapplicabilityforevaluatingotherpancreaticcancervaccine approaches.